By John R. Crowther
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Extra info for Elisa: Theory and Practice (Methods in Molecular Biology)
9. 4. Indirect I-Ag + W Ab + W Antigen Competition Anti-Ab*E + S + Read W +Ai3 Stage (i): Passive adsorption of antigen. Stage (ii): Wash. Stage (iii): Simultaneous incubation of free antigen (test sample): with antibody directed against antigen on plastic at pretitrated dilution. Stage (iv): Wash. Stage (v): Addition of enzyme-labeled antibody against Ab. Stage (vi): Wash. Stage (vii): Addition of substrate. Stage (viii): Read. Competition ELBA 49 Pre-titration Competition-addition of indirect system of samples containing antibodies?
Figure 4 shows the scheme diagrammatically. 2. Indirect I-Ab + w Ag + w AB Sandwich + w Anti- AB *E + W S + Read Stage (i): Passive adsorption of Ab. Stage (ii): Wash. Stage (iii): Addition of antigen. Stage (iv): Wash. Stage (v) Addition of antibody from different species vs antigen. Stage (vi): Wash. Stage (vii): Addition of enzyme-labeled antispecies (directed against AB). 42 Basic Principles of ELISA Fig. 4. Sandwich ELISA-direct. This systemexploits the antibodies attachedto the solid phase to capture antigen.
In this case, a very important epitope might be detected by the MAb, so that the labeling of a single antibody might be justified and form the basis of other assays as described below. 2. Direct-Labeled I-Ab + W Ag*E + Antigen S + Read W Stage (i): Passive adsorption of antibody to plate. Stage (ii): Wash. Stage(iii): Addition and incubation of enzyme-labeledantigen. Direct ELISA 37 Fig. 1. Direct ELISA. Antigen is attached to the solid phase. After washing, enzyme-labeled antibodies are added.